1. Testosterone, Aggression, and Impulsivity in Rats Erik Manke March 7, 2014

2. http://www.youtube.com/watch?v=Yn5iIF95Qho Steroids vs. Water Bottle Arnold Visits Hans and Franz http://screen.yahoo.com/pumping-hans-franz- arnold-schwarzenegger-000000067.html

3. Defining Impulsivity Kerman et al. 2011 – Marked decreases in behavioral/emotional control, lack of perception, brash decision making Batrinos 2012 – Lack of restraint, disuse of PFC, absence of inhibition, and emotionality Wood et al. 2013 – Immediate disregard in decision making, impatience, short term decisions, and reactive aggression

4. Testosterone and Impulsivity Kerman et al. 2011 – bHR rats (impulsive phenotype)  Increased Aggression  Increased Testosterone x2 and Corticosterone Batrinos 2012 – Testosterone  Activated Amygdala  Increased Emotional Activity  Decreased Pre-frontal Inhibition of Motor Control Wood et al. 2013 – Testosterone  Increased Aggression but Reduced CPu TH  Decrease Impulsivity

5. Questions/Themes/Hypotheses Kerman et al. 2011 Hypothesis- bHR rats = higher aggression, altered 5-HTergic cells in brainstem Compare/contrast bHR/bLR rats’ behaviors, neurochemistry, and hormone levels Differential expression of Tph2 and Sert versus c-fos in rats depending on brain region and phenotype

6. Questions/Themes/Hypotheses (cont.) Batrinos 2012 Relationship of Testosterone and Cortisol Testosterone  Activated Amygdala  Increased Emotional Activity  Decreased Pre-frontal Inhibition of Motor Control (increased impulsivity) Cortisol  Increased Pre-frontal cortex control (decreased impulsivity) Seratonin  Inhibits motor impulsivity Testosterone, cortisol, and serotonin form a triad

7. Questions/Themes/Hypotheses (cont.) Wood et al. 2013 How do AAS affect impulsivity? Through DA? – Acb, CPu, PFC, and VTA/SN AAS  Aggressive Behavior  DA from Hypothalamus  Increased aggression and Impulsivity

8. Kerman et al. 2011 High/Low Responder Rats

9. bHR and bLR Rats Selectively-bred high and low responder rats Many generations bred for distinct behaviors bHR rats = Phenotypically impulsive – “…heightened novelty-induced exploration, impulsivity, and increased sensitivity to drugs of abuse.” bLR rats = Phenotypically non-impulsive – “…exaggerated depressive and anxiety-like behaviors.”

10. Serotonin Key role in aggressive responses Influx upon resident/intruder experiments Measured by Tph2 (synthesis) and Sert (reuptake) gene expression Expression inhibits c-fos expression Expect high Tph2 and Sert in bHR rats Expect high c-fos in bLR rats

11. Figure 1: Behavioral differences between bHR and bLR rats

12. Figure 2: Relative Testosterone/Corticosterone Levels in bHR versus bLR rats before and after intrusion

13. Figure 3: Serotonergic cell groups in rat brainstem sections caudal (A) to rostral (T)

14. Figure 4: Tph2 (top) and Sert (bottom) expression differences between bHR rats (left) and bLR rats (right)- Significance in B9 cell group and pontomesencephalic reticular formation

15. Figure 5: Greater Tph2 expression in bHR rats compared to bLR rats

16. Figure 6: Greater Sert expression in bHR rats compared to bLR rats

17. Figure 7: Sert expression (red) and c-fos expresssion (green) and overlay in bLR (left) versus bHR (right)

18. Figure 8: c-fos expression greater in bLR rats compared to bHR rats in certain regions

19. Batrinos 2012 Testosterone and (He?)Man I Said Hey!

20. The triad Testosterone vs. Cortisol/Serotonin PET and fMRI allow locality and interactions to be determined Ratios determine aggressiveness, anti-social behavior, anger, and possibly impulsiveness

21. Testosterone Associated with aggression/anti-social behavior Violent vs. non-violent prisoners Testosterone dosing CAG repeats in human androgen receptor promoter Testosterone  Amygdala  Reduced pre- frontal cortex inhibition (higher impulsivity) Local brain testosterone > effect than circulatory

22. Cortisol Antagonist to testosterone Inhibits GnRH Linked to submissive behavior Cortisol  Testosterone  Decreased Impulsivity Testosterone inhibits CRH Testosterone/Cortisol ratio may predict impulsivity

23. Serotonin Counteracts testosterone Regulates impulsivity and aggressiveness Both activating and inhibitory neurons in pre- frontal and subcortical areas High pre-frontal serotonin = low impulsivity

25. Wood et al. 2013 ‘Roid rage in rats?

26. Nose Poke Test Male rats trained for nose poke response to light for potential fight Resident/Intruder Model (5 min) FI10 Schedule Measure operant response Measure acts of aggression

27. Figure 1: Operant Responses - Significant differences= * -No difference between testo/vehicle for operant responses/rate -Testo rats fought more/earlier -Vehicle rats in contact more often/longer

28. Delayed-Discounting Procedure 2 retractable levers with control on sides Light stimulus 70 s trial with 10 s response window Initially equal rewards 1 forced trial Large reward delay increased by 15 s increments Impulsive = immediate reward (1 pellet) Not Impulsive = delayed reward (4 pellets)

29. Figure 2: Delay-Discounting Impulsivity -No significant differences between Vehicle/Testo Rats -Tested body weight (A) -Food per session (B) -Food per day (C) -Unreinforced Trials per session (D)

30. Figure 3: Large Reward Preference -Small/Immediate reward= impulsive preference -Large/Delayed reward=non- impulsive preference -Only significant at 45 second delay -Trend -Q: Why are testosterone rats less impulsive? A: Look at Immunoblots

31. Western Immunoblot 20 week old rat brains Measure target protein (TH) levels – PFC, Acb, CPu, VTA/SN Primary Antibodies for TH and beta-tubulin Secondary Antibodies for fluorescence Ratio of TH to beta tubulin measured

32. Tyrosine Pathway TH  Rate limiting enzyme

33. Figure 4: Western Immunoblot (Top) and TH Levels (Bottom) -Top-Caudate/Putamen TH and beta tubulin protein expression -Bottom- Testo/Vehicle TH levels only significantly differ in the Caudate/Putamen -Q: What does this mean? A: CPu causes disinhibition of thalamus increasing impulsivity

35. Reactive vs Proactive Aggression Chris Benoit 2007 Double Murder Suicide

36. Answering the Focal Questions Kerman et al. 2011 Impulsive phenotype (bHR) Increase in Aggression  increase 5-HT Two fold increase in testosterone and corticosterone Increased Tph2 and Sert expression Decreased c-fos expression bHR rats exhibit elevated gene expression levels causing elevated aggression (possibly impulsivity?)

37. Answering the Focal Questions (cont.) Batrinos et al. 2012 Testosterone/cortisol ratio and serotonin levels form impulsiveness regulatory triad Primary interaction = amygdala and pre- frontal cortex Aggression and Impulsiveness closely tied

38. Answering the Focal Questions (cont.) Wood et al. 2013 TH levels in Acb, VTA/SN, and PFC = same in Immunoblot study for vehicle/testo CPu TH lower and delayed reward higher in testo reflected lower impulsivity Testosterone does not  Greater impulsivity Testosterone may  Less impulsivity

39. Final Thoughts/Conclusions Three modes of thinking regarding testosterone and impulsivity: 1. Phenotypic impulsivity  higher aggression  higher testosterone 2. Increased testosterone  decrease pre-frontal cortex motor control  increase impulsivity 3. Testosterone  Increased Aggression but Reduced CPu TH  Decrease Impulsivity

40. Works Cited Batrinos, M. L. 2012. Testosterone and aggressive behavior in man. International Journal of Endocrinology & Metabolism 10(3): 563-568. Kerman, I. A., Clinton, S. M., Bedrosian, T. A., Abraham, A. D., Rosenthal, D. T., Akil, H., & Watson, S. J. 2011. High novelty-seeking predicts aggression and gene expression differences within defined serotonergic cell groups. Brain Research 1419: 34-45. Montoya, E. R., Terberg, D., Bos, P. A., & Van Honk, J. 2011. Testosterone, cortisol, and serotonin as key regulators of social aggression: A review and theoretical perspective. Motivation and Emotion 36: 65-73. Wood, R. I., Armstrong, A., Fridkin, V., Shah, V., Najafi, A., & Jakowec, M. 2013. ‘Roid rage in rats? Testosterone effects on aggressive motivation, impulsivity and tyrosine hydroxylase. Physiology & Behavior 110-111: 6-12.